A rat basophil leukemic cell line (RBL-2H3) exhibited phosphorylation of both the 200,000 and 20,000-dalton chains of myosin following antigenic stimulation. Cells were primed with specific IgE and labeled with 32Porthophoshpate and then stimulated with DNP-bovine albumin. Cells were then disrupted and myosin was selectively removed by immunoprecipitation with anti-platelet myosin antibodies. Scanning of the heavy chains in Coomassie-blue stained polyacrylamide gels revealed 1.1 Mug of myosin per 106 cells. Cells, stimulated at 30 degree C for 0.5 to 5 minute, released increasing amounts of histamine, 32% of the total cellular histamine being release in 5 minute. Scanning of autoradiograms suggested that the amount of radioactive phosphate associated with the myosin heavy and light chains increased 2-3 fold over that present in nonstimulated controls, during the same time period. Two- dimensional maps of tryptic digests indicated that the light chains were phosphorylated by myosin light chain kinase, and protein kinase C. Whether this prosphorylation of myosin within intact basophils plays a role in the secretory process, is under investigation.